PROJECT SUMMARY/ABSTRACT Identification of the molecular regulatory points of exercise benefits is of high national priority because of the opportunity to develop targeted novel therapeutics benefiting populations suffering from inactivity-related health problems, including T2DM and pre-diabetes, characterized by insulin resistance (IR). IR is most prevalent in the older population associated with sarcopenia. We propose novel metabolic regulatory role of PGC-1?4 (?4), a hypertrophy gene, enhanced by resistance exercise (RE). Based on substantial preliminary data, we hypothesize that ?4, in cooperation with PPAR? (R?), promotes muscle glycolysis and insulin sensitivity (IS) as well as increasing muscle mass and performance. Based on our novel preliminary data we will also investigate whether by deacetylation of glycolytic proteins, RE enhances muscle glycolytic capacity. R? also reduces oxidative stress that not only enhances IS but also contributes to other health benefits. New mRNA based data indicates that RE reduces protein degradation which will be investigated in the current proposal. We will determine whether 3 months of RE training enhances insulin sensitivity and muscle performance and mass in IR people through pathways of enhanced glycolysis, deacetylation of glycolytic proteins reducing protein degradation and enhancing synthesis and ameliorating oxidative stress. We will study 48 IR people 50-75 yrs before and after 3 months of either 4-times/week resistance training or sedentary life and compare them with lean IS people. We will collect vastus lateralis muscle biopsy samples before and after an acute exercise bout and following a mixed meal to measure markers of glycolysis, energy metabolites, glycogen synthase, glycogen content, ?4, R?, insulin signaling proteins and proteome analysis. We will also measure markers of oxidative stress including 8-OXO-dg (measure of DNA damage), oxidative damage to proteins and subsequent muscle protein degradation, which we hypothesize is reduced by increased anti-oxidant effect of R? with RE training. We also will use in vivo labeling of specific muscle proteins utilizing stable isotope labeled tracers to determine whether ?4 induced muscle hypertrophy occurs not only by reducing degradation but also by enhancing contractile protein synthesis. Although our preliminary cell line studies provide supporting data on direct effects of ?4 and R? on IS and glycolysis and on the anti-oxidant effect of R?, direct effects of these genes on our outcomes cannot be obtained in humans. Therefore we will perform studies in a mouse model with high-fat diet-induced IR to show that ?4 enhances IS and glycolysis and R? reduces oxidative stress. We also will silence ?4 and R? of mouse muscle to confirm our cell based results showing that contraction-induced changes are dependent on ?4 and R?. Together these human and animal studies will render the necessary mechanistic explanation on how RE enhances IS, glycolysis, reduces oxidative stress and promote muscle performance and mass in IR people, thus substantially contributing to their health and life spans.